Large-scale identification of proteins in human salivary proteome by liquid chromatography/mass spectrometry and two-dimensional gel electrophoresis-mass spectrometry
Large-scale identification of proteins in human salivary proteome by liquid chromatography/mass spectrometry and two-dimensional gel electrophoresis-mass spectrometry
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DOI：
10.1002/pmic.200401037
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发表时间：
2005-04-01
期刊：
影响因子：
3.4
通讯作者：
Wong, DT
中科院分区：
文献类型：
--
作者：
Hu, S;Xie, YM;Wong, DT
人的唾液含有大量的蛋白质和多肽(唾液蛋白质组)，有助于维持口腔内的动态平衡。人类唾液蛋白质组的全球分析对于了解口腔健康和疾病发病机制具有重要意义。本研究利用鸟枪式蛋白质组学和双向凝胶电泳质谱(2-DE-MS)对唾液蛋白质进行了大规模鉴定。对于鸟枪法，根据分子量对整个唾液蛋白进行预分提。最小的一部分，推测含有唾液肽，直接用毛细管液相色谱(LC)分离。然而，大的蛋白质组分被消化成多肽，用于随后的LC分离。利用四极飞行时间质谱仪对分离的多肽进行在线电喷雾串联质谱仪(MS/MS)分析，并对获得的光谱进行自动处理，搜索人类蛋白质序列数据库进行蛋白质鉴定。此外，还使用双向凝胶电泳法对唾液中的蛋白质进行了分析。提取105个蛋白质斑点，凝胶内消化，用基质辅助激光解吸/电离-质谱仪测定多肽片段，并用LC-MS/MS进行蛋白质序列分析。总体而言，我们使用这两种蛋白质组学方法对人类全唾液中的309种蛋白质进行了分类。
Human saliva contains a large number of proteins and peptides (salivary proteome) that help maintain homeostasis in the oral cavity. Global analysis of human salivary proteome is important for understanding oral health and disease pathogenesis. in this study, large-scale identification of salivary proteins was demonstrated by using shotgun proteomics and two-dimensinal gel electrophoresis-mass spectrometry (2-DE-MS). For the shotgun approach, whole saliva proteins were prefractionated according to molecular weight. The smallest fraction, presumably containing salivary peptides, was directly separated by capillary liquid chromatography (LC). However, the large protein fractions were digested into peptides for subsequent LC separation. Separated peptides were analyzed by on-line electrospray tandem mass spectrometry (MS/MS) using a quadrupole-time of flight mass spectrometer, and the obtained spectra were automatically processed to search human protein sequence database for protein identification. Additionally, 2-DE was used to map out the proteins in whole saliva. Protein spots 105 in number were excised and in-gel digested; and the resulting peptide fragments were measured by matrix-assisted laser desorption/ionization-mass spectrometry and sequenced by LC-MS/MS for protein identification. In total, we cataloged 309 proteins from human whole saliva by using these two proteomic approaches.